The master transcriptional regulator Runx2 is essential for osteogenesis and has multifunctional molecular and biological. Runx2 is properties as a scaffolding protein that interacts with distinct co-regulatory factors and is targeted by a unique Runx2 nuclear matrix targeting signal to subnuclear domains. Runx2 is highly expressed in tumor cells that metastasize to bone, present at trace levels in non-metastatic malignant cells and nearly absent from normal mammary epithelial cells. We have proven that the unique targeting function of Runx2 is an essential for its activity which promotes tumor growth in bone and osteolytic bone disease. We have shown that Runx2 is a transcriptional activator of many genes involved in early and late events of metastasis and mediates signaling pathways that contribute to tumor growth. Therefore, we hypothesize that Runx2 regulates a cohort of genes abnormally activated or repressed genes in highly metastatic breast cancer cells in subnuclear domains of tumor cells in mammary gland that will promote metastasis to distal sites. Importantly, we have established that loss of Runx2 fundion in tumor cells (by Runx2 shRNAs and a subnuclear targeting deficient (STD) mutation) reduces tumor growth in the mammary gland and blocks metastatic bone disease. Thus, we propose to further understand the mechanisms of Runx2 activities that are responsive to the mammary tumor microenvironment and are aberrantly associated with subnuclear foci in tumor cells. Our aims are to 1- Establish that Runx2 promotes metastasis of breast cancer cells from primary mammary tumors and that disruption of Runx2 in nuclear microenvironments will decrease metastatic events in a genetic mouse model; 2- Characterize the specific Runx2 subnudear-dependent functions that are required for tumor growth in the mammary fat pad and the bone microenvironment; and 3- Identify specific regulatory proteins in subnuclear domains of breast cancer cells that support to tumor growth and metastasis. These studies, in collaboration with Projects 1 and 2, will define Runx2 as a principal mediator of tumor growth and metastasis by identifying novel Runx2-dependent signaling pathways which function in nuclear microenvironments and contribute to progression of breast cancer.